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    Identity of Epicoccum sorghinum Causing Leaf Spot Disease of Bletilla striata in China

     

    论文题目

    Identity of Epicoccum sorghinum Causing Leaf Spot Disease of Bletilla striata in China

    作    者

    H.Zhou(周浩),P.P.Liu,S,Qiu,S.J.Wei,K.Xia,Q.Gao

    发表期刊

    Plant Disease

    //

    2018102(5)

    摘    要

    Bletilla striata (Thunb.) Rchb. f. (Orchidaceae), known as a traditional Chinese herb, has been widely used as hemostatic and anti-microbial. In May 2017, a leaf spot disease was observed on 20–30% of B. striata in commercial fields located in Ziyuan County, Guangxi, China. Small, circular, brown spots were early observed on leaves, enlarging and developing into irregular dark brown and necrotic lesions. Necrotic lesions caused leaf abscission and occurred frequently during the plant’s life cycle. Three symptomatic leaves from three individual plants were surface sterilized in 1% sodium hypochlorite for 2 min, rinsed 3 times in sterile water,plated on potato dextrose agar (PDA) medium and then incubated at 26°C for 3 days.Seven isolates recovered from diseased plants and the colonies were villose and regular, pale grey, secreting scarlet pigment to the media. After 15 days of incubation, subspheroical chlamydospores were observed, which were brown to black, with verrucose surface. Unicellular chlamydospores were 12.9 to 29.3 × 13.5 to 32.7 μm while multicellular chlamydospores were 6.2 to 13.6 × 29.7 to 89.4 μm. Fungal colonies grown under ultraviolet light for 12 hour with a distance of 60 cm produced numerous pycnidia and conidia on PDA. Brown pycnidia ranged from 90 to 240 μm in diameter. Conidia were hyaline, ellipsoidal, unicellular, aseptate, 5.0 to 7.1 × 2.9 to 3.7 μm in size. Based on the above morphological characteristics, these isolates were identified as Epicoccum sp. (previously as Phoma, Yuan et al. 2016). The identity of isolate was confirmed by sequence analysis of rDNA-ITS, TUB and ACT of strain BJ-F1 (GenBank MF948994, MF987525 and MF987526, respectively). The primers of ITS, TUB and ACT were respectively ITS1/ITS4, T1/Bt2b,ACT-512F/ACT-783R (White et al. 1990; Glass et al. 1995; O'Donnell et al. 1997; Ignazio et al.1999). BLAST results showed that the rDNA-ITS gene sequence was 99% to those of E. sorghinum (= P. sorghina) in GenBank (KJ767080 and KY454466), while the TUB and ACT gene sequences was even 100% homologous to those of the above species (KU728638 and KU728639). Further molecular phylogenetic analysis by Maximum Likelihood method based on the combined ITS-TUB-ACT sequences revealed that strain BJ-F1 was closest to E. sorghinum(previously as E. sorghi, Aveskamp et al. 2010). Pathogenicity tests were conducted on six young plants (ca. 1.5 years old) by inoculating unwounded leaves with three, 5 × 5 mm mycelial discs of two isolates on PDA. Three control plants were inoculated with sterile PDA discs. All inoculated and control plants were enclosed in transparent plastic bags and incubated in a greenhouse at 26°C for 13 days with a photoperiod of 12 h. The first lesions appeared 3 days after inoculation on leaves, and typical symptoms developed 10 days later. Uninoculated controls remained asymptomatic. The experiments were repeated 3 times with similar results. To fulfill Koch's postulates, the fungus was re-isolated from the inoculated leaves on PDA, and confirmed as E.Page 1 of 4 Plant Disease "First Look" paper .

     


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